Methods for the treatment of erectile dysfunction by human birth tissue material composition

ABSTRACT

A composition for treating erectile dysfunction is provided. The composition includes a human birth tissue material composition for topical application or introduction onto or into a penis affected by erectile dysfunction.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a divisional application of U.S. patent applicationSer. No. 14/215,765, filed on Mar. 17, 2014, which claims priority toU.S. Provisional Application No. 61/802,428 filed Mar. 16, 2013, each ofwhich is incorporated by reference herein in its entirety.

FIELD OF THE INVENTION

Methods for treating erectile dysfunction by administering a human birthtissue material composition are provided.

BACKGROUND OF THE INVENTION

Erectile Dysfunction (ED) is a repeated inability to achieve or maintaina penile erection for sexual intercourse. It is estimated that about 30million men in the United States and 150 million men worldwideexperience chronic erectile dysfunction. Additionally, studies haveshown that about half of the men between the ages of 40 and 70 haveerectile dysfunction to some degree. The prevalence of erectiledysfunction is illustrated by the fact that Viagra, an ED drugmanufactured by Pfizer, has been prescribed to more than 35 million menworldwide as of 2011.

The human penis consists of three erectile bodies: two corpora cavernosachambers and one corpus spongiosum chamber. Together these threechambers make up the expandable erectile tissues along the length of thepenis which fill with blood during penile erection. The two corporacavernosa lie along the penis shaft, from the pubic bones to the head ofthe penis, where they join. These formations are made of a sponge-liketissue containing irregular blood-filled spaces lined by endothelium andseparated by trabeculae of smooth muscle fibers with an extracellularmatrix of which the main components are collagens, elastic fibers,proteoglycans/glycoseaminoglycans, in addition to numerous unmyelinatedand preterminal autonomic nerves.

The importance of corporal cavernosal smooth muscle cells (SMCs) inpotency is well-established. Normal smooth muscle content and functionare necessary for the initiation and maintenance of erection.Additionally, elastin and collagen fibers are also important penileconstituents. Collagen is a key structural protein in tissues subjectingto stretching forces. Collagen provides structural integrity that allowsthe cavernosum to withstand pressure increase during erection. Someresearchers argue that without this rigid collagen network, the peniswould maintain a flaccid form.

Erectile dysfunction can be caused by a number of factors, includingphysical, psychological, neurological, vascular and endocrinal issues.Reduced blood flow to the penis and nerve damage are the most commoncauses of erectile dysfunction. Underlying causes include, but are notlimited to, vascular disease, diabetes, drugs, hormone imbalance,neurological causes, pelvic trauma in surgery, Peyronie's Disease andvenous leak.

Currently, the most common method for treating erectile dysfunction isthrough oral inhibitors of phosphodiesterase V (PDE-5), including themedications: Sildenafil (Viagra®), Vardenafil (Lavetra®) and Tadalafil(Cialis®). However, numerous patients remain unresponsive to treatment,do not tolerate the adverse effects associated with the treatment, orare ineligible for treatment. Thus, there remains a need for a safe andeffective treatment of erectile dysfunction which can be used withoutthe side effects associated with the use of PDE-5 inhibitors.

SUMMARY OF THE INVENTION

A method of treating erectile dysfunction in a subject is provided. Themethod includes the step of administering a human birth tissue materialcomposition onto or into an affected penis. The birth tissue materialcomposition comprises one or more of the components of the placentalorgan, including but not limited to, the placental globe, the umbilicalcord, the umbilical cord blood, the chorionic membrane, the amnioticmembrane, the Wharton's jelly, the amniotic fluid, and other placentalgelatins, cells and extracellular material. The administration caninclude topical application, urethral suppository, intracavernousinjection or a combination thereof.

According to another aspect, a kit for treating erectile dysfunction isprovided that includes the composition as provided herein. The kit mayalso include at least one delivery device. The kit may further comprisea set of instructions for use thereof.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a schematic providing an overview of the method of preparing ahuman birth tissue material composition according to one embodiment;

FIG. 2 provides a batch volume and fill calculation sheet according toone embodiment; and

FIG. 3 provides a solution calculation sheet according to oneembodiment.

DETAILED DESCRIPTION OF THE INVENTION

The present disclosure will now be described more fully hereinafter withreference to exemplary embodiments thereof. These exemplary embodimentsare described so that this disclosure will be thorough and complete, andwill fully convey the scope of the disclosure to those skilled in theart. Indeed, the present disclosure may be embodied in many differentforms and should not be construed as limited to the embodiments setforth herein; rather, these embodiments are provided so that thisdisclosure will satisfy applicable legal requirements. As used in thespecification, and in the appended claims, the singular forms “a”, “an”,“the”, include plural referents unless the context clearly dictatesotherwise.

As used herein, “human birth tissue” encompasses one or more of thecomponents of the placental organ including, but not limited to, theplacental globe, the umbilical cord, the umbilical cord blood, thechorionic membrane, the amniotic membrane, the Wharton's jelly, theamniotic fluid, and other placental gelatins, cells, and extracellularmaterial.

As used herein, “placental tissue components” encompass one or more ofthe tissue components of the placental organ including, but not limitedto, the placental globe, the umbilical cord, the umbilical cord blood,the chorionic membrane, the amniotic membrane, the Wharton's jelly andother placental gelatins, cells and extracellular material.

As used herein, the term “amnion” and “amniotic membrane” are usedinterchangeably.

As used herein, the term “effective amount” refers to an amount of aparticular composition sufficient to elicit the desired therapeuticeffects.

A method of treating erectile dysfunction in a subject is provided. Themethod includes the step of administering a human birth tissue materialcomposition onto or into an affected penis. The birth tissue materialcomposition comprises one or more of the components of the placentalorgan, including but not limited to, the placental globe, the umbilicalcord, the umbilical cord blood, the chorionic membrane, the amnioticmembrane, the Wharton's jelly, the amniotic fluid, and other placentalgelatins, cells and extracellular material. The administration caninclude topical application, urethral suppository, intracavernousinjection or a combination thereof.

The birth tissue material composition administered in the methods of thepresent invention may be prepared as provided herein. One embodiment ofa method for preparing a birth tissue material composition is providedin the schematic of FIG. 1. The method of preparing a human birth tissuematerial composition includes the step of recovering placental tissuecomponents and amniotic fluid from a seronegative, healthy human.Potential human birth tissue donors providing informed consent arepre-screened during an examination of pre-natal medical records andblood test results. A comprehensive medical history and behavior riskassessment is obtained from the donor prior to donation incorporatingU.S. Public Health Service guidelines. Discussions with the physician(s)and/or the donor mother are conducted to identify circumstances that maylead to the exclusion of the donor or donated tissue. Additionally, aphysical exam is performed on the donor to determine whether there isevidence of high risk behavior or infection and to determine the overallgeneral health of the donor.

Infectious disease testing of donor blood specimens is performed foreach tissue donor on a specimen collected at the time of donation orwithin seven days prior to or after donation. Advantageously, themethods that are used to screen for a communicable disease follow theregulations as set forth by the Federal Drug Administration and theAmerican Association of Tissue Banks. Exemplary infectious diseasetesting includes, but is not limited to, antibodies to the humanimmunodeficiency virus, type 1 and type 2 (anti-HIV-1 and anti-HIV-2);nucleic acid test (NAT) for HIV-1; hepatitis B surface antigen (HBsAg);total antibodies to hepatitis B core antigen (anti-HBc-total, meaningIgG and IgM); antibodies to the hepatitis C virus (anti-HCV); NAT forHCV; antibodies to human T-lymphotropic virus type I and type II(anti-HTLV-I and anti-HTLV-II); and syphilis (a non-treponemal ortreponemal-specific assay may be performed).

Human birth tissue is preferably recovered from a full-term Cesareandelivery of a newborn. Alternatively, human birth tissue is recoveredfrom a full-term vaginal delivery of a newborn. The subsequent steps ofpreparing the human birth tissue material are performed in a controlledenvironment (i.e., certified biological safety cabinet, hood or cleanroom). Instruments, solutions, and supplies coming into contact with thehuman birth tissue material during processing are sterile. All surfacescoming in contact with the human birth tissue material intended fortransplant are either sterile or draped using aseptic technique.

Once recovered, one or more of the placental tissue components can beremoved via a sterile saline solution rinse, blunt dissection, scalpel,or a combination thereof, if necessary. According to one embodiment, theplacental globe, umbilical cord, chorionic membrane, and other gelatins,fluids, cells and extracellular matrix are removed and discarded,leaving the amniotic membrane for further processing. In a preferredembodiment, the human birth tissue material is subject to the method ofpreparation described herein no more than four hours after recovery topreserve cell viability.

The retained placental tissue components can be placed in a steriletransport solution after aseptic recovery. The sterile transportsolution is used to provide an advantageous medium to the naturalfunction of the placental tissue components prior to processing. Forexample, calcium-rich water can be used as the sterile transportsolution to provide a medium to drive undifferentiated cells to becomeosteogenic when implanted. Throughout the preparation of the human birthtissue material, various methods can be used to drive undifferentiatedcells to differentiate into specialized cell types including, but notlimited to, transport solutions, soaks, particular temperature ranges,and hyperbaric pressure.

The sterile transport solution preferably includes sodium chloride(NaCl) in a concentration range from typically about 10% to typicallyabout 20% by weight. The sterile transport solution can also include oneor more of Minimum Essential Medium, Dulbecco's Modified Eagle's Medium,Plasma Lyte-A, human albumin 25% solution, calcium-rich water, alkalineionized water, or acidic ionized water.

Amniotic fluid can be placed in a sterile container after asepticrecovery. In a preferred embodiment, a cell count is performed on theamniotic fluid using methods commonly known in the art (e.g.,hemocytometer). The amniotic fluid is preferably mixed thoroughly beforea sample is taken to ensure that the sample is representative of thenumber of cells dispersed throughout the amniotic fluid. Additionally,an appropriate dilution of the mixture with regard to the number ofcells to be counted can be utilized. The total cell count per millilitercan then be calculated. In another embodiment, a cell counter can beused to determine total cell count per milliliter of fluid. After thecell count is determined, a requisite cell suspension solution volumecan be calculated and prepared. The cell suspension volume may becalculated from predetermined requirements for the minimum starting gramweight of tissue per mL of bulk tissue product. In an alternateembodiment, the cell suspension volume may be calculated frompredetermined requirements for the minimum number of amniotic fluidcells per 1.0 mL aliquot of bulk tissue product. In one embodiment, thecell suspension solution includes typically about 91% volume of PlasmaLyte-A and typically about 9% volume of human albumin 25% solution.

In a preferred embodiment, after the cell count is completed, anamniotic fluid composition is prepared according to the following steps.Particles in the amniotic fluid are separated from the liquid componentof the amniotic fluid using centrifugation. The resulting, separatedparticles are referred to as “pellets.” The separation of particles fromthe liquid component of the amniotic fluid may occur by anyart-recognized method including sedimentation or microfiltration. In apreferred embodiment, the amniotic fluid is evenly aliquoted intosterile conical centrifuge tubes. The amniotic fluid can be distributedin equal amounts in as many tubes as necessary for the volume recovered.The amniotic fluid can be centrifuged at 200 rpm to 15,000 rpm for aperiod of up to 30 minutes at ambient temperature. In one embodiment,the amniotic fluid is centrifuged at approximately 1410 rpm (400×gravity[RCF]) for a period of 10 minutes at ambient temperature. Using asterile pipette, the supernatant can be aspired from each tube anddiscarded. An isotonic solution can be used to re-suspend each pelletand bring the volume of each tube up to a predetermined amount. In oneembodiment, the isotonic solution is Plasma Lyte-A. The pellet/isotonicsolution combination can be centrifuged at 200 rpm to 15,000 rpm for aperiod of up to 30 minutes at ambient temperature. In one embodiment,the pellet/isotonic solution combination is centrifuged at approximately1410 rpm (400×g [RCF]) for a period of 10 minutes at ambienttemperature. Using a sterile pipette, the second supernatant can beaspired from each tube and discarded. Each second pellet can bere-suspended in a cell suspension solution of a predetermined amount toform the amniotic fluid composition. In one embodiment, the cellsuspension solution includes Plasma Lyte-A and human albumin 25%solution. In a preferred embodiment, the cell suspension solutionincludes typically about 91% volume of Plasma Lyte-A and typically about9% volume of human albumin 25% solution. The cells in each tube can bethoroughly suspended by using a vortex mixer for a minimum of threeseconds. Immediately thereafter, the contents of each tube arehomogenized with a cell suspension solution to form the amniotic fluidcomposition. In a preferred embodiment, the cell suspension solutionincludes typically about 91% volume of Plasma Lyte-A and typically about9% volume of human albumin 25% solution.

In one embodiment, the discarded first and second supernatant from theaforementioned amniotic fluid composition preparation steps are furtherprecipitated using dialysis equipment or micropore/nucleopore filters.Alternately, the first and second supernatant can be furtherprecipitated by pipetting the supernatants onto sterile wax paper,heating the contents to quickly evaporate the liquid, and then addingthe remaining material back into the final amniotic fluid composition.This method allows for the removal of the extraneous liquid, whilemaximizing the cells, proteins and other particles otherwise discardedin the first and second supernatants. The remaining materialprecipitated from the first and second supernatants can be homogenizedwith the amniotic fluid composition.

After the amniotic fluid composition is generated, a second cell countcan be performed on a representative sample using a hemocytometer, acell counter, or any other method commonly known in the art. Theamniotic fluid preparation can be stored at typically about 1-10° C. fora period of up to 24 hours pending further processing.

A predetermined percentage of the amniotic fluid composition(representative sample) can be retained for testing and not included inthe final bulk product. This representative sample can be retained foranalysis and comparison to the cells in the final bulk product todiscern any deleterious effects on the amniotic fluid cells,particularly the effects of the cryoprotectant(s) on the amniotic fluidcells in the final bulk product and/or the effects of cleaning,processing, freezing and thawing on the amniotic fluid cells.

Simultaneous with the initial cell count of the amniotic fluid, theweight of the placental tissue components can be determined. Thereafter,the placental tissue components can be transferred aseptically to asterile dish containing Plasma Lyte-A and stored in a quarantinerefrigerator pending further processing.

After the weight of the placental tissue components is determined, arequisite tissue suspension solution volume can be calculated andprepared based on predetermined requirements for the minimum startinggram weight of tissue per mL of bulk tissue product. In one embodiment,the tissue suspension solution comprises Plasma Lyte-A, human albumin25% solution, and dimethyl sulfoxide. In a preferred embodiment, thetissue suspension solution comprises: typically about 44% volume ofPlasma Lyte-A, typically about 36% volume of human albumin 25% solution,and typically about 20% volume of dimethyl sulfoxide.

In one embodiment, the total number of final product units can becalculated based on predetermined requirements for: (1) the minimumstarting tissue gram weight per mL of bulk tissue product; and (2) theminimum number of amniotic fluid cells per 1.0 mL aliquot of bulk tissueproduct. In a preferred embodiment, the total number of final productunits can be calculated based on predetermined requirements for theminimum starting tissue gram weight per mL of bulk tissue product. Inone embodiment, the final product vials can be of various volumes suchas, for example, 0.25 mL, 0.50 mL, 1.0 mL, 1.25 mL, 2.0 mL, 3.0 mL orany other volume as contemplated by one of ordinary skill in the art.

In one embodiment, the placental tissue components include amnioticmembrane. In another embodiment, the placental tissue components includeone or more components selected from the group consisting of amnioticmembrane, chorionic membrane, Wharton's jelly, umbilical cord tissue,umbilical cord blood, placental globe, and other gelatins, other cellsand extracellular matrix from placental tissue components. The placentaltissue components can then be placed in a sterile dish containing PlasmaLyte-A until further processing.

The placental tissue components can be removed from the Plasma Lyte-Aand cryopreserved according to methods commonly used in the art. Theplacental tissue components can be soaked in cryoprotectant prior tocryopreservation. In one embodiment, the cryoprotectant is one commonlyused in the industry, such as, for example, dimethyl sulfoxide (DMSO).In a preferred embodiment, the cryoprotectant is an amnion control ratefreeze solution comprising typically about 44% volume of Plasma Lyte-A,typically about 36% volume of human albumin 25% solution, and typicallyabout 20% volume of dimethyl sulfoxide. In another embodiment, thecryoprotectant is a commercially available cryoprotectant such asSynth-a-Freeze® available from Invitrogen. Any cryoprotectant specificto the birth tissue material described herein may be used. In oneembodiment, cryopreservation is achieved using a controlled ratefreezer, resulting in a 1° C. rate from nucleation to −35° C. and a 10°C. per minute cooling rate to a −90° C. end temperature. However, anycryopreservation method commonly known in the art may be used.

After cryopreservation, the placental tissue components are subjected tomorselization. As used herein, “morselization” means to grind up toparticle form. Tissue morselization may occur by any art-recognizedmethod of tissue disruption, including, but not limited to: milling,blending, sonicating, homogenizing, micronizing, pulverizing,macerating, or a combination thereof. In one embodiment, the placentaltissue components are subjected to cryogenic milling by methods commonlyknown in the art. In a preferred embodiment, the tissue is cryogenicallymilled in a CryoMill® (available from Retsch) for two cycles at afrequency 1/s of 25 Hz with a pre-cooling time of no more than aboutfive minutes, a grinding time of no more than about two minutes, and anintermediate cooling time of no more than about five minutes. In anotherembodiment, a Freezer/Mill® available from SPEX SamplePrep, LLC may beused. In one embodiment, the total number of final product units can becalculated based on predetermined requirements for: (1) the minimumtissue gram weight after morselization per mL of bulk tissue product;and (2) the minimum number of amniotic fluid cells per 1.0 mL aliquot ofbulk tissue product. In an alternate embodiment, the total number offinal product units can be calculated based on predeterminedrequirements for the minimum tissue gram weight after morselization permL of bulk tissue product.

After morselization, the milled placental tissue components can becombined with the tissue suspension solution to form a tissuesuspension. In one embodiment, the tissue suspension solution includesPlasma Lyte-A, human albumin 25% solution, and dimethyl sulfoxide, whichis used immediately to prepare the final bulk tissue product. In apreferred embodiment, the tissue suspension solution comprises typicallyabout 44% volume of Plasma Lyte-A, typically about 36% volume of humanalbumin 25% solution, and typically about 20% volume of dimethylsulfoxide, which is used immediately to prepare the final bulk tissueproduct. In an alternate embodiment, the tissue suspension solutionincludes typically about 44% volume of Plasma Lyte-A and typically about36% volume of human albumin 25% solution. The 20% volume of dimethylsulfoxide is purposefully withheld pending final combination of the bulktissue product. In this alternate embodiment, the milled tissuesuspension (without dimethyl sulfoxide) can be stored at about 1-10° C.for a period of up to about 24 hours, pending further processing. In thealternate embodiment, the 20% volume of dimethyl sulfoxide can be addedto the tissue suspension immediately prior to final bulk tissue productmanufacture.

Bulk tissue product can be manufactured by homogenizing the amnioticfluid composition and the tissue suspension. Both the amniotic fluidcomposition and the tissue suspension can be vortexed for no less thanabout three seconds prior to combination. In a preferred embodiment, thebulk tissue product can be homogenized using a laboratory homogenizermixer, followed by continuous mixing with magnetic stirrers. Immediatelythereafter, the bulk tissue product can be placed on cold packs andindividual, empty cryovials can be filled with the bulk tissue product.In one embodiment, the final product vials can be of various volumessuch as, for example, 0.25 mL, 0.50 mL, 1.0 mL, 1.25 mL, 2.0 mL, 3.0 mLor any other volume as contemplated by one of ordinary skill in the art.Cryopreservation of the final bulk tissue product vials can be achievedthrough control-rate freezing by methods commonly known in the art.

Representative samples from the beginning, middle, and end of thecryovial fill cycle can be removed from the final product count forquality control testing, including, but not limited to, bioburdentesting, mycoplasma DNA by polymerase chain reaction, and bacterialendotoxin test (Limulus Ameboycte Lysate). Representative samples fromthe beginning, middle, and end of the cryovial fill cycle can be removedfrom the final product count to store for future testing should the needarise.

Another embodiment of a method for preparing a birth tissue materialcomposition is provided herein. According to one embodiment, thecomposition includes human placental tissue components. The compositioncan optionally include an acceptable carrier composition. The humanplacental tissue components can be prepared according to the stepsprovided herein. In certain embodiments, the human placental tissuecomponents can be combined with the carrier composition to formulate acomposition suitable for application onto or into a penis affected witherectile dysfunction.

To prepare the human placental tissue components for inclusion in acomposition, placental tissue components are initially recovered from aseronegative, healthy woman per the aforementioned donor screeningprocedures, including a comprehensive medical history and behavior riskassessment, physical exam and infectious disease testing.

Placental tissue is preferably recovered from a full-term Cesareandelivery of a newborn. Alternatively, placental tissue is recovered froma full-term vaginal delivery of a newborn. The subsequent steps ofpreparing the placental tissue components are performed in a controlledenvironment (i.e., certified biological safety cabinet, hood or cleanroom). Instruments, solutions, and supplies coming into contact with thehuman placental tissue material during processing are sterile. Allsurfaces coming in contact with the human placental tissue materialintended for transplant are either sterile or draped using aseptictechnique.

According to one embodiment, the human placental tissue components caninclude one or more components selected from the group consisting ofamniotic membrane, chorionic membrane, Wharton's jelly, umbilical cordtissue, umbilical cord blood, placental globe, and other gelatins, othercells and extracellular matrix from placental tissue. Other variationsof the invention include, however, removing one or more of the amnioticmembrane, chorionic membrane, Wharton's jelly, umbilical cord tissue,umbilical cord blood, placental globe, and other gelatins, other cellsand extracellular matrix from placental tissue before furtherprocessing. In a preferred embodiment, the placental tissue componentsinclude amniotic membrane only. Removal of one or more of the placentaltissue components can be achieved via a sterile saline solution rinse,blunt dissection, scalpel, or a combination thereof, if necessary.

The retained placental tissue components can be placed in a steriletransport solution after aseptic recovery. The sterile transportsolution is used to provide an advantageous medium to the naturalfunction of the placental tissue components prior to processing.Throughout the preparation of the human placental tissue composition,various methods can be used to drive undifferentiated cells todifferentiate into specialized cell types including, but not limited to,transport solutions, soaks, particular temperature ranges, andhyperbaric pressure.

The sterile transport solution preferably includes sodium chloride(NaCl) in a concentration range from typically about 0.9% to typicallyabout 20% by weight. The sterile transport solution can also include oneor more of Minimum Essential Medium, Dulbecco's Modified Eagle's Medium,Plasma Lyte-A, human albumin 25% solution, calcium-rich water, alkalineionized water, or acidic ionized water.

After delivery to the processing facility, the weight of the placentaltissue components can be determined. Thereafter, the placental tissuecomponents can be transferred aseptically to a sterile dish containingPlasma Lyte-A and stored in a quarantine refrigerator pending furtherprocessing.

The placental tissue components can be removed from the Plasma Lyte-Aand cryopreserved according to methods commonly used in the art. Theplacental tissue components can be soaked in cryoprotectant prior tocryopreservation. Various cyroprotectants are often used to avoid theformation of ice crystals and/or delay the onset of ice formation to thelowest temperature possible. Such cryoprotectants known in the art aretypically glycerol, dimethyl sulfoxide (DMSO), dimethyl acetamide,methanol, ethylene glycol, propylene glycol, trimethylamine acetate, andother high molecular weight solutes capable of forming strong hydrogenbonds to water, which may be used singularly or in combination thereof.In one embodiment, the cryoprotectant is dimethyl sulfoxide (DMSO) at aconcentration of about 10% (v/v). In a preferred embodiment, thecryoprotectant is an amnion control rate freeze solution comprisingPlasma Lyte-A, human albumin 25% solution, and dimethyl sulfoxide. Inanother embodiment, the cryoprotectant is a commercially availablecryoprotectant such as Synth-a-Freeze® available from Invitrogen.However, any cryoprotectant specific to the placental tissue componentsdescribed herein may be used. In one embodiment, cryopreservation isachieved using a controlled rate freezer, resulting in a 1° C. rate fromnucleation to −35° C. and a 10° C. per minute cooling rate to a −90° C.end temperature. However, any cryopreservation method commonly known inthe art may be used.

According to one embodiment, after cryopreservation, the placentaltissue components are subjected to morselization. As used herein,“morselization” means to grind up to particle form. Tissue morselizationmay occur by any art-recognized method of tissue disruption, including,but not limited to: milling, blending, sonicating, homogenizing,micronizing, pulverizing, macerating, or a combination thereof. In oneembodiment, the placental tissue components are subjected to cryogenicmilling by methods commonly known in the art. In a preferred embodiment,the tissue is cryogenically milled in a CryoMill® (available fromRetsch) for two cycles at a frequency 1/s of 25 Hz with a pre-coolingtime of no more than about five minutes, a grinding time of no more thanabout two minutes, and an intermediate cooling time of no more thanabout five minutes. In another embodiment, a Freezer/Mill® availablefrom SPEX SamplePrep, LLC may be used.

After morselization, the milled placental tissue components can behomogenized with a tissue suspension solution to form the final product.In one embodiment, the tissue suspension solution includes PlasmaLyte-A, human albumin 25% solution, and dimethyl sulfoxide. In apreferred embodiment, the tissue suspension solution comprises typicallyabout 67.5% volume of Plasma Lyte-A, typically about 22.5% volume ofhuman albumin 25% solution, and typically about 10% volume of dimethylsulfoxide. In another embodiment, the tissue suspension solutioncomprises typically about 0.01%-10% volume of dimethyl sulfoxidecombined with a medium or buffer, including, but not limited to, sodiumchloride, glycerol, methylcellulose or appropriate buffer solutions,e.g., phosphate buffered saline.

Final product can be manufactured by homogenizing the placental tissuecomponents and the tissue suspension solution. In a preferredembodiment, the final product can be homogenized using a laboratoryhomogenizer mixer, followed by continuous mixing with magnetic stirrers.Immediately thereafter, the final product can be placed on cold packsand aliquoted into individual, empty cryovials. In one embodiment, thefinal product can be of various volumes such as, for example, 0.25 mL,0.50 mL, 1.0 mL, 1.25 mL, 2.0 mL, 3.0 mL or any other volume ascontemplated by one of ordinary skill in the art. Cryopreservation ofthe final bulk tissue product vials can be achieved through control-ratefreezing by methods commonly known in the art.

Representative samples from the beginning, middle, and end of thecryovial fill cycle can be removed from the final product count forquality control testing, including, but not limited to, bioburdentesting, mycoplasma DNA by polymerase chain reaction, and bacterialendotoxin test (Limulus Ameboycte Lysate). Representative samples fromthe beginning, middle, and end of the cryovial fill cycle can be removedfrom the final product count to store for future testing should the needarise.

In one embodiment, the total number of final product units can becalculated based on predetermined requirements for the minimum startingtissue gram weight per mL of final product. In an alternate embodiment,the total number of final product units can be calculated based onpredetermined requirements for the minimum tissue gram weight aftermorselization per mL of final product.

Another embodiment of a method for preparing a birth tissue materialcomposition is provided herein. According to one embodiment, thecomposition includes human placental tissue components. In some aspects,the placental tissue components may be subjected to morselizationfollowing recovery. As used herein, “morselization” means to grind up toparticle form. Tissue morselization may occur by any art-recognizedmethod of tissue disruption, including, but not limited to, milling,blending, sonicating, homogenizing, micronizing, pressing, pulverizing,triturating, macerating, or a combination thereof. Particles may bemicron or submicron size. The resulting product may be a dry powder. Insome embodiments, the morselized placental tissue components aredissolved in one or more biocompatible solvents to create a paste,emulsion, suspension or solution. This dissolution may occur during themanufacturing process or immediately prior to application. Examples ofbiocompatible solvents include, but are not limited to, physiologicalsaline; BSS™, a balanced salt solution containing per ml 0.64% sodiumchloride, 0.075% potassium chloride, 0.048% calcium chloride, 0.03%magnesium chloride, 0.39% sodium acetate, and 0.17% sodium citratedihydrate, as well as sodium hydroxide and/or hydrochloric acid toadjust pH, and water; Ocular Irrigation Solution™; Lactated Ringer'ssolution; normal saline solution; or normal saline adjusted to pH 7.4with sodium bicarbonate.

In some embodiments, the placental tissue components are morselized byuse of a tissue grinder (e.g., a Potter-Elvehjem grinder or a WheatonOverhead Stirrer). In some embodiments, the placental tissue componentsare micronized by use of a sonicator. In some embodiments, the placentaltissue components are micronized by use of a bead beater. In someembodiments, the placental tissue components are micronized by use of afreezer/mill (e.g., a SPEX SamplePrep Freezer/Mill). In someembodiments, the placental tissue components are micronized by manualuse of a pestle and mortar. The placental tissue components may beoptionally lyophilized before or after being subjected to micronization.

The human birth tissue material compositions as described herein can beoptionally mixed with bioactive agents such as physiologicallycompatible minerals, growth factors, wound healing agents (e.g.,cytokines including but not limited to PDGF, TGF, and thymosin),hyaluronic acid, wound sealants (such as fibrin with or withoutthrombin), cellular attractant and scaffolding reagents (e.g.,fibronectin) antibiotics, chemotherapeutic agents, antigens, antibodies,enzymes, vectors for gene delivery and hormones.

The human birth tissue compositions as described herein can beoptionally mixed with a suitable carrier to form a composition suitablefor treatment of an affected penis from erectile dysfunction. Thecarrier can include any variety of components suitable for applicationonto or into an affected penis. According to one embodiment, the carriercomposition includes one or more vitamins, minerals, proteins, fats,collagens (including collagen extracted from the placental globe),hyaluronic acid, waxes, glycols and derivatives thereof, glyercols andderivatives thereof, oils (including essential oils), fatty acids,cholesterols, alcohols, emollients, adsorbents, lubricants, emulsifyingagents, thickening agents, humectants, surfactants, pharmaceuticalingredients, preservatives, antifungal agents, antioxidants,antimicrobial agents, structuring agents, dispersing agents,pH-adjusting components, sequestering or chelating agents, wettingagents, coloring agents, and other components known in the art to besuitable for use in a composition that can be applied onto or within thehuman body. The optional carrier composition can be formulated in such away that the combination of the human birth tissue material compositionand the carrier composition are chemically compatible and do not formcomplexes which precipitate from the final composition.

According to one embodiment, the human birth tissue material compositionand the carrier composition as provided herein can be mixed or blendedaccording to a variety of conventional techniques. According to oneembodiment, the human birth tissue material composition and the carriercomposition can be mixed in a manner to produce a smooth and homogenouscomposition. According to one embodiment, the human birth tissuematerial composition as provided herein is introduced to the carriercomposition after the carrier composition is formed (i.e., post-added).In an alternative embodiment, the human birth tissue materialcomposition is introduced during carrier composition preparation. Theamount of placental tissue present in the composition can vary dependingupon whether a carrier composition is utilized, the frequency of use,and the treatment desired. The amount of carrier composition present inthe final composition can vary according to the final formulation of thecomposition. According to one embodiment, the carrier compositioncomponents can be present in an amount from typically about 0.1% toabout 99.0% based on total composition weight.

According to a preferred embodiment, the human birth tissue materialcomposition may be administered in combination with other agents knownto increase erectile function, including, but not limited to, inhibitorsof PDE-5, PGE-1, papaverine, promorphine, or other known vasodilators.In another aspect of the invention, the human birth tissue materialcomposition may be administered in combination with testosteronetherapy.

Methods of treating erectile dysfunction are provided. According to oneembodiment, the method includes the step of administering an effectiveamount of a human birth tissue material composition onto or into anaffected penis. A single application or multiple applications may beadministered either to a single site or to more than one site on oraround the penis. Multiple applications may occur essentially at thesame time or separated in time.

According to a preferred embodiment, the composition can be formulatedinto an injectable formulation. An injectable formulation can includethe placental tissue compositions as described herein either alone or incombination with a suitable carrier. The injectable formulation can beadministered via urethral suppository or intracavernous injection.

According to another preferred embodiment, the composition can beapplied topically on or around the penis. Such a topical formation caninclude a placental tissue composition as described herein either aloneor in combination with a suitable carrier composition. According to suchan embodiment, the composition can be formulated as solid, semi-solid,or liquid. Suitable formulations may include, but are not limited to, acream, emulsion, spray, gel, ointment, salve, butter, gel, putty, balm,or pliable stick. In one embodiment, the gel or putty carrier could beachieved through collagen extracted from the placental globe.

Also provided is a kit for treating erectile dysfunction that includesan effective amount of a composition as described herein. Such kits caninclude a package that is adapted to receive one or more containers, atleast one of the container(s) including a composition as describedherein. Containers can include a bottle, a vial (i.e., injectable form),a metal tube, a laminate tube, a plastic tube, a dispenser, apressurized container, or other types of containers such as glass orinjection or blow-molded plastic containers. The containers can dispensea predetermined amount of the composition. In other embodiments, thecontainer can be squeezed (e.g., metal, laminate, or plastic tube) todispense a desired amount of the composition. The containers can havespray, pump, or squeeze mechanisms. In certain embodiments, thecontainer can be a vial that contains sufficient volume of compositionfor one (i.e., single use) or more applications or injections. The kitis appropriately preserved up until and during shipment to adistributor, medical facility, or other end user. According to apreferred embodiment, the kit may include at least one delivery device.The kit can additionally include at least one set of instructions forthe end user, including an explanation of how to apply, inject, andotherwise use and maintain the composition.

Although specific embodiments of the present invention are hereinillustrated and described in detail, the invention is not limitedthereto. The above detailed descriptions are provided as exemplary ofthe present invention and should not be construed as constituting anylimitation of the invention. Modifications will be obvious to thoseskilled in the art, and all modifications that do not depart from thespirit of the invention are intended to be included with the scope ofthe appended claims.

Having generally described the present invention, a furtherunderstanding can be obtained by reference to the examples providedherein for purposes of illustration only and are not intended to belimiting.

Example 1

The placental construct may be prepared according to the method of FIG.1, the details of which are herein provided.

Human birth tissue was obtained from a seronegative, healthy mother viaCesarean section. To maximize the overall quality of the donated tissue,a recovery technician was present in the operating room during thedonor's Cesarean section to assist the surgical team with recovery,treatment and handling of the birth tissue. The donor was surgicallyprepped and draped per AORN standards prior to the Cesarean sectionprocedure. The recovery technician prepared the recovery site byestablishing a sterile field on a back table in the operating room.

Amniotic fluid was recovered according to the following proceduresprovided herein. The physician's assistant cleared all maternal bloodfrom the surgical site. A suction cannula was positioned directly abovethe intended amnion/chorion membrane incision site. Using the smallestappropriate incision, the amniotic and chorionic membranes werebreached, releasing the amniotic fluid into the suction cannula.Avoiding maternal blood, the physician's assistant suctioned as muchamniotic fluid volume as possible into a sterile suction canister.Immediately following recovery, the sterile suction canister wastransferred to the sterile back table. The recovery technician examinedthe amniotic fluid for the presence of visible blood. After noting thatno visible blood was present, the recovery technician asepticallytransferred the amniotic fluid to a sterile Nalgene jar and performedswab cultures. The recovery technician secured the lid on the Nalgenejar to which the appropriate identification was affixed.

Following delivery of the baby, the physician's assistant placed thehuman birth tissue en-bloc into a sterile basin. Maintaining sterility,the basin was transferred to the recovery technician onto the sterilefield on the back table. Beginning at the amnion/chorion membranesurgical incision site, the recovery technician used blunt dissection toseparate the chorionic membrane from the amniotic membrane, using carenot to tear the amniotic membrane. The recovery technician then removedthe amniotic membrane from the placental face until reaching theumbilical cord. At the site where the amnion is attached to theumbilical cord, the recovery technician dissected the amnion from theumbilical cord by making an incision in the amnion around thecircumference of the umbilical cord. The amniotic membrane wastransferred to a sterile container and rinsed with sterile saline toremove any blood or debris.

After thorough rinsing, the amniotic membrane was transferred into asterile bag and swab cultures were performed. Approximately 300 mL oftransport solution (15% NaCl) was added to the sterile bag containingthe recovered amniotic membrane. The bag was secured with a knot. Thesingle-bagged amniotic membrane was then placed into a second sterilebag, which was securely knotted. The double-bagged amniotic membrane wasthen transferred into a plastic transport container to which theappropriate identification was affixed.

The Nalgene jar containing the amniotic fluid and the plastic transportcontainer containing the amniotic membrane were placed in a qualifiedshipping container with an appropriate amount of wet ice to maintainrefrigerated temperatures. The validated box arrived at the processingfacility approximately one hour following recovery and was immediatelyinspected per standard operating procedures and placed in refrigeratedtemperatures (1-10° C.).

Processing was performed on a sterile field using aseptic technique in avalidated cleanroom at the processing facility less than four hoursafter the recovery was completed. All manufacturing steps were recordedconcurrently by a circulating technician on a designated processingrecord.

Amniotic Membrane Preparation

The amniotic membrane was removed from the plastic transport containerand outer bag. The inner bag containing the amniotic membrane wasaseptically passed onto a sterile field. Approximately 35 mL of thesterile transport solution was aspirated out of the bag utilizing asterile pipette. Subsequently, the sample was transferred to a sterileconical tube for pre-processing bioburden testing. Using sterileforceps, the amniotic membrane was removed from the inner bag and placedin a sterile basin on a sterilely draped pre-set balance. The weight ofthe amniotic membrane was recorded. The sterile basin containing theamniotic membrane was transferred to a back table. Approximately 250 mLof Plasma Lyte-A was added to a second sterile basin and covered withthe corresponding sterile lid. Using sterile forceps, the amnioticmembrane was removed from the first sterile basin and transferred to asterile prep board where the membrane was spread flat. Using a sterilelap sponge, any remaining debris/blood was removed from the surface ofboth sides of the amniotic membrane. The amniotic membrane wassubsequently transferred to a second sterile basin containing PlasmaLyte-A where the membrane was covered, labeled and transferred to aquarantine refrigerator.

Amniotic Fluid Preparation

Sterile pipettes and 50 mL sterile conical centrifuge tubes weretransferred to a sterile field. The Nalgene jar was moved in a gentleswirling motion to ensure cells were equally dispersed throughoutamniotic fluid prior to removal of samples. The Nalgene jar containingthe amniotic fluid was opened, and 10 mL of amniotic fluid was aspiratedout utilizing a sterile pipette and transferred to a sterile conicaltube for pre-processing bioburden testing. Approximately one mL ofamniotic fluid was aspirated out utilizing a sterile pipette in order tocomplete the cell count. Utilizing a sterile 50 mL pipette, theremaining amniotic fluid was aseptically aspirated out of the Nalgenejar and transferred into 50 mL conical centrifuge tubes. Aliquots of theamniotic fluid were aseptically distributed in equal amounts in an evennumber of 50 mL sterile conical centrifuge tubes.

Batch Volume and Aliquot Fill Calculation

The batch volume and aliquot fill calculations (See FIG. 2) weredetermined based on the following calculations.

-   -   1. Record Amnion Weight (AW) in grams.    -   2. Calculate Amnion Allowable Aliquots (AA).    -   2.1 Divide Amnion Weight (AW) by minimum starting amnion gram        weight per 1 mL aliquot (0.03 grams) to calculate total Amnion        Allowable Aliquots (AA).    -   3. Calculate Cell Count        -   3.1 Record the total Amniotic Fluid Volume (A) in mL.        -   3.2 Record the Total Cells Counted (B) for the four large            corner squares and the middle square of the hemocytometer            counting grid using the hemocytometer and an inversion            microscope per standard operating procedures.        -   3.3 Calculate Average Cells/Square (C).            -   3.3.1 (C)=Total Cells Counted (B)/5 Squares Counted        -   3.4 Record Dilution Factor used in preparation of cell count            (D).        -   3.5 Calculate the Total Cell Density (cells/mL) (E).            -   3.5.1 (E)=(C)×(D)×10⁴        -   3.6 Calculate Total Cells (TC).            -   3.6.1 Total Cells (TC)=Total Cell Density (E)×Total                Volume of Amniotic Fluid (A)    -   4. Calculate Aliquot Cell Density (CD) (Aliquot=1 mL).        -   4.1 (CD) mL=Total Cell (TC)/Amnion Allowable Aliquot (AA)    -   5. Determine Bulk Product Volume (BV).        -   5.1 (AA) Amnion Allowable Aliquots=Bulk Product Volume (BV)    -   6. Determine Lot Vial Fill Calculations.        -   6.1 Record Bulk Product Volume (BV)        -   6.2 Record the target vial production per size provided by            management.        -   6.3 Calculate Total Vial Target        -   6.4 Calculate Total Volume Requirement for Vial Target.            -   6.4.1 [Vial Target for 0.25 mL vials]×0.25=Volume                Requirement (mL) for 0.25 mL Vials            -   6.4.2 [Vial Target for 0.50 mL vials]×0.50=Volume                Requirement (mL) for 0.5 mL Vials            -   6.4.3 [Vial Target for 1.25 mL vials]×1.25=Volume                Requirement (mL) for 1.25 mL Vials        -   6.5 Calculate Total Vial Fill Volume.        -   6.6 Compare Bulk Product Volume to Total Vial Fill Volume            requirement based on the production plan.            -   6.6.1 Total Vial Fill Volume must be Bulk Product Volume                (BV).            -   6.6.2 Adjust vial targets accordingly.

Solutions Calculations

After obtaining the Bulk Product Volume (BV), the component volumes forthe tissue suspension solution (i.e., amnion suspension solution) andthe cell suspension solution were determined based on the followingcalculations (See FIG. 3).

1.0 Bulk Product Volume (mL) (BV)=Total Suspension Solution Volume (mL)(SS)

2.0 Calculate Cell Suspension Solution Volume (CS)

-   -   2.1 (CS)=Total Suspension Solution Volume (SS)/2    -   2.2 Calculate (CS) Component Volume(s) Required:        -   2.2.1 Plasma Lyte-A Volume (mL)=(CS)×0.91        -   2.2.2 Human Albumin 25% Solution Volume (mL)=(CS)×0.09    -   2.3 Calculate Amnion Suspension Solution (AS)        -   2.3.1 (AS)=Total Suspension Solution Volume (SS)/2        -   2.3.2 Calculate (AS) Component Volume(s) Required:            -   2.3.2.1 Plasma Lyte-A Volume (mL)=(AS)×0.44            -   2.3.2.2 Human Albumin 25% Sol. Volume (mL)=(AS)×0.36            -   2.3.2.3 Dimethyl Sulfoxide, USP Volume (mL)=(AS)×0.20

Solution Preparations

The following materials were transferred to the sterile field: (i) HumanAlbumin 25% Solution, Excipient, EU Grade; (ii) Plasma Lyte-A Injection(pH 7.4); and (iii) DMSO (dimethyl sulfoxide), USP. In separate 1 Lsterile containers, the cell suspension solution and the amnionsuspension solution were prepared based on the calculations obtainedutilizing the solution calculations sheet set forth in FIG. 3. Theamnion control rate freeze solution was prepared according to thedirections as set forth in FIG. 3. The containers were labeled withrespective solution names, lot numbers, and expiration dates and storedat 1-10° C. pending further use.

Aseptic Processing of Amniotic Fluid Composition

Amniotic fluid-filled conical tubes were aseptically transferred to anEppendorf centrifuge and centrifuged at 400×g for 10 minutes at ambienttemperature. At the completion of each cycle, the conical tubes wereaseptically transferred back to the sterile field. Each conicalcentrifuge tube was checked by a processing technician to ensure apellet had formed. The results were recorded in the batch record. Thesupernatant was removed and discarded using a sterile pipette, and asufficient volume of Plasma Lyte-A was added to each conical tube tore-suspend the pellet and increase the volume in each tube toapproximately 20 mL. Each tube was placed on a vortex mixer for 3-5seconds to fully re-suspend the pellets. The contents of the conicalcentrifuge tubes were subsequently combined, reducing the overallconical centrifuge tube number by half by quickly pouring the suspensionfrom a first tube to a second tube, ensuring maximum transfer of cellsduring combination. The process was repeated until all remaining conicalcentrifuge tubes were combined, reducing the number of tubes by half.The remaining conical centrifuge tubes were aseptically transferred toan Eppendorf centrifuge and centrifuged at 400×g for 10 minutes atambient temperature. At the completion of each cycle, the conical tubeswere aseptically transferred back to the sterile field. Each conicalcentrifuge tube was checked by a processing technician to ensure apellet had formed. The results were recorded in the batch record. Thesupernatant was removed and discarded using a sterile pipette, and asufficient volume of cell suspension solution was added to each conicaltube to re-suspend the pellet and increase the volume in each tube toapproximately 20 mL. Each tube was placed on a vortex mixer for 3-5seconds to fully re-suspend the pellets. Next, each suspension wasquickly poured into the container of cell suspension solution to formthe amniotic fluid composition. The amniotic fluid composition wasstored in refrigerated temperatures at 1-10° C. until furtherprocessing.

Amnion Control Rate Freezing

The following materials were transferred to the sterile field: amnion(in Plasma Lyte-A solution); amnion control rate freeze solution;appropriately sized pipettes; sterile bowl; sterile forceps, steriletray; and sterile spatula. The amnion was removed from Plasma Lyte-Asolution and transferred to the amnion control rate freeze solution.After 30 minutes, the amnion was removed from the amnion control ratefreeze solution and transferred to the sterile tray where it was cutinto four equal sections. The tray with the sectioned amnion wasaseptically transferred to a control rate freezer. A control ratefreezer probe was placed near the center of the chamber, taking care notto contact any metal in the chamber. The control rate freezer wasactivated by selecting a pre-programmed cycle.

Amnion Morselization

The amnion was subjected to morselization by cryogenic milling by theprocedures described herein. A Spex Freezer/Mill® was programmed to thefollowing settings: grinding rate=12; cycles=3; pre-cooling time=5minutes; grinding time=2 minutes; and intermediate cooling=2 minutes.Sterile, autoclaved milling cylinders, impactors and end-caps wereplaced in an ultra-low freezer for a minimum of 15 minutes in order topre-cool the materials prior to use. The milling cylinders, impactor andend-caps were removed from ultra-low freezer and aseptically transferredto the sterile field. One end cap was inserted onto each cylinder. Theamnion was subsequently removed from the control rate freezer. Oneamnion section was placed into each of the four cylinders. An impactorbar was placed inside each of the four cylinders. The second end-cap wassecured onto each cylinder, sealing the four milling chambers. Themilling chambers were placed into the Spex Freezer/Mill® one at a timeand allowed to run for the aforementioned program settings. At theconclusion of each milling event, the chamber was removed andimmediately aseptically transferred to a sterile field. Using a sterileextractor tool, an end cap was removed from each chamber. The impactorbar and milled amnion were quickly dispensed into a sterile bowl. Asterile spatula was used to remove any remaining milled amnion from themilling cylinder or end-caps. Approximately 100 mL of amnion suspensionsolution was added to the milled amnion in a sterile bowl. Once thawed,any remaining amnion was removed from the impactor using a sterilespatula. This milling procedure was repeated for each of the fourmilling chambers until all milled amnion was added to the amnionsuspension solution, thereby forming the tissue suspension.

Bulk Tissue Product

A sterile 2 L Erlenmeyer flask was aseptically transferred to a backtable. The tissue suspension (amnion suspended in the amnion suspensionsolution) and the amniotic fluid composition were aseptically pouredinto the 2 L Erlenmeyer flask. The flask was appropriately covered andlabeled. Immediately thereafter, the flask was placed in a quarantinerefrigerator at 1-10° C.

Vial Fill

The following materials were transferred to the sterile field: sterilepipettes; sterile cryovial racks; sterile cryovials; and bulk tissueproduct. The bulk tissue product was removed from the quarantinerefrigerator and placed on cold packs on a sterilely draped mixer. Astir bar was aseptically added to the bulk tissue product. The cryovialswere filled using a repeater pipette pre-set to target fill volume asindicated in the production plan. Immediately following fill andcapping, each cryovial was inspected per quality control (QC) standardoperating procedures. Any vials failing QC inspection were discarded perbiohazard waste disposal standard operating procedures. The cryovialsthat passed QC inspection were placed in cryovial racks.

Bulk Tissue Product Cryopreservation

The cryovial racks were transferred to sterile racks and placed in acontrol rate freezer. A control rate freezer probe was placed near thecenter of the chamber, taking care not to contact any metal in thechamber. The control rate freezer was activated by selecting apre-programmed cycle. Upon completion of each control rate freeze, eachcryovial was inspected per QC standard operating procedures. Any vialsfailing QC inspection were discarded per biohazard waste disposalstandard operating procedures. The cryovial racks were placed in sterilecontainers and transferred to a quarantine ultra-low freezer to awaitresults of all lot release testing before final packaging.Representative samples from the beginning, middle, and end of thecryovial fill cycle were removed from the final product count for lotrelease testing, which included: bioburden testing, mycoplasma DNA bypolymerase chain reaction, and bacterial endotoxin test (LimulusAmeboycte Lysate).

Packaging of Cryopreserved Bulk Tissue Product

Throughout packaging procedures, the cryovials containing bulk tissueproduct were exposed to ambient temperature for a time period of oneminute or less. After lot release testing clearance, each cryovial waspackaged into a sterile foil pouch using aseptic technique. Using anAccuSeal 540Plus sealer, each foil pouch was sealed following standardoperating procedures. Following QC inspection, each pouch was packagedin an outer box and labeled with the unique tissue identification numberassigned to the cryovial, which was designed to ensure the traceabilityof the tissue from receipt through clinical use, transfer ordestruction. Each cryovial was stored at ultra-low temperatures (≤−65°C.) prior to transplantation. Final product vial sizes were 0.25 mL,0.50 mL, and 1.25 mL.

Example 2 (Prophetic)

Five (5) adult male subjects with organic erectile dysfunction ofgreater than one (1) year due to nerve-sparing radical prostatectomy,diabetes mellitus and/or vascular disease will participate in a six (6)month prospective, nonrandomized clinical feasibility study. The studysubjects had scores of less than 26 on a validated, diagnosticquestionnaire, II EF-EF (Erectile Function Domain of the InternationalIndex of Erectile Function).

The human birth tissue material composition is substantially preparedaccording to the method of FIG. 1 (see Example 1). Each study subjectwill receive up to 30 cc of the composition injected into the corporacavernosum, with one (1) injection on the base of each side of thepenis, for a total of two (2) injections over a two (2) to four (4)minute period. A compression band will be applied to the base of thepenis for 10-15 minutes during the entirety of the procedure. Given thefact that 30 cc of solution is needed to create an erection (ensuresthat all cavernosal tissue is affected), the injection medium mayinclude the human birth tissue material composition alone or thecomposition homogenized with a sterile saline.

Initial safety will be assessed by examination of adverse eventsreported immediately after treatment at the following intervals: 24hours, 48 hours, 72 hours, one (1) week and two (2) weeks. A summaryregarding safety will be prepared by the Principal Investigator aftercompletion of the study, including acute and long-term adverse events.

Efficacy will be assessed based on assessments at pre-treatment(baseline), six (6) weeks, three (3) months and (6) monthspost-treatment using validated, diagnostic questionnaires andself-report diaries. The primary objective of the of the efficacyassessments is to evaluate the ability of the human birth tissuematerial composition to restore erectile function at six (6) monthsusing II EF-EF scores. However, secondary objectives include: (i)summarizing the single item Erection Hardness Score (EHS) at baselineand at each follow-up visit; (ii) summarizing the penile length (flaccidand stretched) and girth at baseline and at each follow-up visit; and(iii) summarizing the number of sexual intercourse attempts andpercentage of subjects with positive responses to Sexual EncounterProfile Question 2 (SEP-2) and

Question 3 (SEP-3) at baseline and at each follow-up visit.

We claim:
 1. A composition for treating erectile dysfunction comprising:a human birth tissue material composition for topical application orintroduction onto or into a penis affected by erectile dysfunction, thehuman birth tissue material composition comprising a therapeuticallyeffective amount of a one or more components of a placental organselected from the group consisting of morselized placental globe,morselized umbilical cord, umbilical cord blood, morselized chorionicmembrane, morselized amniotic membrane, Wharton's jelly, amniotic fluid,and other placental gelatins, cells, and morselized extracellularmaterial, wherein the morselized placental component(s) is/arehomogenized with a tissue suspension solution, the tissue suspensionsolution comprising human albumin 25% solution and dimethyl sulfoxide,and wherein the composition is suitable for restoration of erectilefunction.
 2. The composition of claim 1, wherein the components of theplacental organ are cryopreserved prior to morselization.
 3. Thecomposition of claim 1, wherein the human birth tissue materialcomposition further comprises an agent selected from the groupconsisting of PDE-5 inhibitor, PGE-1, papaverine, promorphine,testosterone or any combination thereof.
 4. The composition of claim 1,wherein the human birth tissue material composition is configured to bedelivered in the form of a topical application, urethral suppository,intracavernous injection, or a combination thereof.
 5. The compositionof claim 4, wherein the topical application is a cream, emulsion, spray,gel, ointment, salve, butter, gel, putty, balm, or pliable stick.
 6. Akit for treating erectile dysfunction comprising an effective amount ofa human birth tissue material composition as provided in claim
 1. 7. Thekit of claim 6, further comprising at least one delivery device.
 8. Thekit of claim 6, further comprising instructions for use thereof.